Figure 10. Reverse transcriptase polymerase chain reaction analysis of markers during retinal pigment epithelium cell differentiation. mRNA was isolated from undifferentiated (A) or differentiated (B) ARPE-19 cells and subjected to reverse transcriptase polymerase chain reaction amplification to detect SMA (C) or RPE65 (D) as described in Methods. Phase-contrast micrographs represent undifferentiated (A) or differentiated (B) ARPE-19 cells after one week (A) or 52 weeks (B) of culture. C and D represent RT–PCR amplification of mRNA samples isolated from ARPE-19 cells maintained in culture for 1, 2, 5, 7, 10, 12, and 52 weeks, using primers to detect mRNA for either αSMA (C) or RPE65 (D). E represents RT–PCR amplification for a series of 25, 30, or 35 cycles to detect αSMA using mRNA isolated from ARPE-19 cells that are undifferentiated (U) differentiated (D). The first lane in C-E represents a DNA standard ladder of 300, 400, and 500 bp.